Physical proximity and functional interplay of the glycoprotein Ib-IX-V complex and the Fc receptor FcgammaRIIA on the platelet plasma membrane.
Although the glycoprotein (GP) Ib-IX-V complex and FcgammaRIIA are distinct platelet membrane receptors, previous studies have suggested that these structures may be co-localized. To determine more directly the proximity of GP Ib-IX-V and FcgammaRIIA, we assessed the effects of anti-GP Ibalpha monoclonal antibodies on FcgammaRIIA-mediated platelet aggregation and on the direct binding of polymeric IgG to human platelets. In addition, we directly examined the proximity of FcgammaRII and GP Ib-IX-V using flow cytometric fluorescence energy transfer and immunoprecipitation studies. Preincubation of platelets with either of two monoclonal antibodies (AN51 or SZ2) directed against GP Ibalpha completely blocked platelet aggregation by polymeric IgG. Similarly, these antibodies totally inhibited platelet aggregation by two strains of viridans group streptococci known to induce aggregation via FcgammaRIIA. In addition, AN51 and SZ2 significantly reduced the binding of polymeric IgG to washed fixed platelets. When assessed by flow cytometry, significant levels of bidirectional energy transfer were detected between FcgammaRIIA and GP Ibalpha, indicating a physical proximity of less than 10 nm between these receptors. This energy transfer was not due to high receptor density, because no homoassociative energy transfer was seen. Moreover, immunoprecipitation of FcgammaRIIA from platelet lysates also co-precipitated GP Ibalpha. These results indicate that GP Ibalpha and FcgammaRIIA are co-localized on the platelet membrane and that this association is not random.
Sullam
P M
PM
Department of Medicine, University of California, San Francisco and Veterans Affairs Medical Center, San Francisco, California 94121, USA. sullam@sanfrancisco.va.gov
Hyun
W C
WC
Szöllösi
J
J
Dong
J f
Jf
Foss
W M
WM
López
J A
JA
D000911:Antibodies, Monoclonal; D015703:Antigens, CD / Q000302:isolation & purification* / Q000378:metabolism; D001779:Blood Coagulation Factors / Q000302:isolation & purification* / Q000378:metabolism; D001792:Blood Platelets / Q000187:drug effects / Q000276:immunology / Q000378:metabolism / Q000648:ultrastructure*; D002462:Cell Membrane / Q000187:drug effects / Q000276:immunology / Q000378:metabolism; D004735:Energy Transfer; D005456:Fluorescent Dyes; D006801:Humans; D007074:Immunoglobulin G / Q000494:pharmacology; D010974:Platelet Aggregation; D019038:Platelet Glycoprotein GPIb-IX Complex / Q000302:isolation & purification* / Q000378:metabolism; D011233:Precipitin Tests; D011485:Protein Binding; D017452:Receptors, IgG / Q000302:isolation & purification* / Q000378:metabolism
D000911:Antibodies, Monoclonal; D015703:Antigens, CD; D001779:Blood Coagulation Factors; C080568:Fc gamma receptor IIA; D005456:Fluorescent Dyes; D007074:Immunoglobulin G; D019038:Platelet Glycoprotein GPIb-IX Complex; D017452:Receptors, IgG; C037685:platelet factor V; C044411:polymeric IgG